U73122

U73122 significantly inhibits aggregation of human platelets induced by a variety of agonists, including collagen, thrombin, ADP, arachidonic acid, with IC50 of 1-5 μM. U-73122 (10 μM) inhibits the production of IP3 and the subsequent rapid increase in cytosolic Ca²⁺ induced by either thrombin or U-46619 through inhibiting hydrolysis of phosphatidyl[³H]inositol and phosphatldyl[³H]inosito1 4,5-bisphosphate catalyzed by a soluble fraction from platelets (K_i=9 and 40 μM, respectively). U-73122 inhibits thromboxane B production induced by collagen through inhibiting receptor-coupled mobilization of arachidonic acid. U73122 inhibits also FMLP-induced aggregation of human polymorphonuclear neutrophils and the associated production of IP3 and diacylglycerol. ^[1]

U-73122 causes a concentration-dependent inhibition of C5a, FMLP, PAF and LTB4-induced MPO and B12-BP release from cyto-chalasin B-treated PMNs. The IC50 values are 60 (FMLP), 110 (LTB4), 115 (C5a) and 120 (PAF) nM for MPO release; and 105 (FMLP), 110 (LTB4), 120 (C5a) and 140 (PAY) nM for B12-BP release. U-73122 is also a potent inhibitor of superoxide anion production by cytochalasin B-treated PMNs activated with either C5a or FMLP with IC50 of 160 and 300 nM, respectively. U-73122 causes suppression of the rise in [Ca²⁺]_i, IP3 production and DAG production in FMLP-stimulated PMNs with IC50 of 500 nM, 2 μM, and 2 μM, respectively. 3 μM U-73122 causes 100% inhibition of FMLP-induced GTPase activity. U-73122 causes a concentration-dependent inhibition of the FMLP-evoked association of PKC with the extractable particulate fraction of PMNs, but not a soluble preparation of PMN PKC. ^[2]

U73122 significantly inhibits recombinant human PLC-β2, with an IC50 of ~6 μM. U73122 has little effect on PLC-β1, PLC-β3, or PLC-β4. U73122 reduces interleukin-8 and leukotriene B4-induced Ca²⁺ flux and chemotaxis in human neutrophils with IC50 of ~6 μM and ~5 μM, respectively. ^[3]

1 μM U73122 blocks bradykinin (BK)-induced increases in the intracellular free Ca²⁺ concentration in undifferentiated NG108-15 cell. The IC50 for a 20-min exposure is approximately 200 nM. 1 μM U73122 produces a small but significant increase in [Ca²⁺]_i which results from Ca ²⁺ release from an intracellular store. In differentiated NG108-15 cells U73122 blocks completely depolarization-induced Ca²⁺ influx. In contrast, in DRG neurons U73122 inhibits only slightly voltage-sensitive Ca²⁺ channels. ^[4]

U73122 is a relatively specific inhibitor of G-protein-mediated phospholipase C activation in pancreatic acini. U73122 inhibits phosphatidylinositol (PI) hydrolysis on stimulation with either cholecystokinin (by 81%) or carbachol (by 73%) at a maximal effect concentration of 10 μM. U73122 (10 μM) inhibits the increases in [Ca²⁺]_i stimulated by high concentrations of secretagogues in fura-2-loaded acini. U73122 also inhibits the [Ca²⁺]_i signal generated by directly stimulating G-proteins with sodium fluoride. U73122 rapidly inhibits the oscillating [Ca²⁺]_i signal elicited by low concentrations of cholecystokinin or carbachol. ^[5]

U73122 increases the activity of hPLCβ3 in a concentration-and time-dependent manner in a cell-free micellar system, with up to an 8-fold increase in enzyme activity and a EC50 of 13.6 μM. Activation of hPLCβ3 by U73122 requires covalent modification of cysteines. U73122 (10 μM) also activates hPLCγ1(>10-fold) and hPLCβ2(~2-fold); PLC δ1 is neither activated nor inhibited. ^[6]

U73122 (30 mg/kg i.p.) blocks swelling of rats hind paw by 65 and 80% at 1 and 3 h postcarrageenan challenge. U73122 (0.1 mg/mL) inhibits carrageenan-induced macrophage and lymphocyte accumulation into subcutaneous chambers in dogs by 65 and 74%, respectively. U73122 (30 mg/kg i.p.) totally inhibits the LPS-induced increase in macrophage and lymphocyte infiltration and prostaglandin E2 production (by 80%) in a mouse peritonitis model, and inhibits and 12- O-tetradecanoyl-phorbol-13-acetate-induced ear edema in mice. ^[3]

• Phosphoinositide-specific phospholipase C activity assay

  Assay mixtures (0.2 mL) contains assay buffer (without leupeptin), CaCl₂ (an amount calculated to produce a Ca-EGTA buffer with the required free Ca²⁺ concentration), platelet soluble fraction (5-15 μg of protein) and 4 nmol of either phosphatidyl[³H]inositol or phosphatidyl[³H]inositol 4,5-bisphosphate. Substrata mixtures consists of either phosphatidyl[³H]inositol (3.4 Ci/mol) plus 1-pahnitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (1:0.4 M ratio) or phosphatidyl[³H]inositol4,5-bisphosphate (7 Ci/mol) plus 1-palmitoyl 2-oleoyl-sn-giycero-3-phosphoethanolamine (1:4 M ratio). Substrate mixtures are prepared as l0× suspensions of small unilamellar vesicles by consonication of the lipids in assay buffer (Vibra Cell microprobe sonicator, 75 W, 2 times 60 sec). U-73122 is added to assay mixtures in 2 μL of DMSO. This amount of DMSO has a negligible effect on phospholipase C activity under these conditions. Assay mixtures are incubated in glass tubes at 37 ℃ for 10 min and then quenched with 1.6 mL of chloroform/rnethanol/HC1 (1 N) (108:108:25, by vol). After vigorous mixing and centrifugation (500 g for 10 min), a portion (0.7 mL) of the aqueous upper layer (0.9 mL) is transferred to a scintillation vial that contains 10 mL of ACS scintillation fluid. Tritium in water-soluble products (>90% being inositol phosphates) is measured by liquid scintillation spectrornetry. Phosphoinositide hydrolysis under these conditions never exceeds 10% of total added and proceeded at a constant rate throughout the incubation period. Blank values are determined from assay mixtures in which platelet soluble fraction is added after the stopping reagent.

• Animal Models

  Swiss-Webster mic

• Dosages

  30 mg/kg

• Administration

  i.p.